美国药典USP31-NF26无菌检查法《71》.doc

71 STERILITY TESTS 无菌检查法

Portions of this general chapter have been harmonized with the corresponding texts of the European Pharmacopeia and/or the Japanese Pharmacopeia. Those portions that are not harmonized are marked with symbols ( ) to specify this fact.

此通则的各部分已经与欧洲药典和/或日本药典的对应部分做了协调。不一致的部分用符号（ ）来标明。

The following procedures are applicable for determining whether a Pharmacopeial article

purporting to be sterile complies with the requirements set forth in the individual monograph with respect to the test for sterility. Pharmacopeial articles are to be tested by the Membrane Filtration method under Test for Sterility of the Product to be Examined where the nature of the product permits. If the membrane filtration technique is unsuitable, use the Direct Inoculation of the

Culture Medium method under Test for Sterility of the Product to be Examined. All devices, with the exception of Devices with Pathways Labeled Sterile, are tested using the Direct Inoculation of the Culture Medium method. Provisions for retesting are included under Observation and Interpretation of Results.

下面这些步骤适用于测定是否某个用于无菌用途的药品是否符合其具体的各论中关于无菌检查的要求。只要其性质许可，这些药品将使用供试产品无菌检查法项下的膜过滤法来检测。如果膜过滤技术是不适合的，则使用在供试产品无菌检查法项下的培养基直接接种法。除了具有标记为无菌通道的设备之外，所有的设备均须使用培养基直接接种法进行检测。在结果的观测与理解项下包含了复验的规定。

Because sterility testing is a very exacting procedure, where asepsis of the procedure must be

ensured for a correct interpretation of results, it is important that personnel be properly trained and qualified. The test for sterility is carried out under aseptic conditions. In order to achieve such conditions, the test environment has to be adapted to the way in which the sterility test is performed. The precautions taken to avoid contamination are such that they do not affect any microorganisms that are to be revealed in the test. The working conditions in which the tests are performed are monitored regularly by appropriate sampling of the working area and by carrying out appropriate controls.

由于无菌检查法是一个非常精确的程序，在此过程中程序的无菌状态必须得到确保以实现对结果的正确理解，因此人员经过适当的培训并取得资质是非常重要的。无菌检查在无菌条件下进行。为了实现这样的条件，试验环境必须调整到适合进行无菌检查的方式。为避免污染而采取的特定预防措施应不会对任何试图在检查中发现的微生物产生影响。通过在工作区域作适当取样并进行适当控制，来定期监测进行此试验的工作条件。

These Pharmacopeial procedures are not by themselves designed to ensure that a batch of product is sterile or has been sterilized. This is accomplished primarily by validation of the sterilization process or of the aseptic processing procedures.

这些药典规定程序自身的设计不能确保一批产品无菌或已经灭菌。这主要是通过灭菌工艺或者无菌操作程序的验证来完成。

When evidence of microbial contamination in the article is obtained by the appropriate

Pharmacopeial method, the result so obtained is conclusive evidence of failure of the article to meet the requirements of the test for sterility, even if a different result is obtained by an alternative procedure. For additional information on sterility testing, see Sterilization and Sterility Assurance of Compendial Articles 1211 .

当通过适当的药典方法获得了某物品中微生物污染的证据，这样获得的结果是该物品未能达

到无菌检验要求的结论性证据，即便使用替代程序得到了不同的结果也无法否定此结果。如要获得关于无菌检验的其他信息，见药品的灭菌和无菌保证<1211>

MEDIA 培养基

Prepare media for the tests as described below, or dehydrated formulations may be used provided that, when reconstituted as directed by the manufacturer or distributor, they meet the requirements of the Growth Promotion Test of Aerobes, Anaerobes, and Fungi. Media are sterilized using a validated process.

按照下面描述的方法配制实验用培养基；或者使用脱水培养基，只要根据其制造商或者分销商说明进行恢复之后，其能够符合好氧菌、厌氧菌、霉菌生长促进试验的要求即可。使用经过验证的工艺对培养基进行灭菌操作。

The following culture media have been found to be suitable for the test for sterility. Fluid

Thioglycollate Medium is primarily intended for the culture of anaerobic bacteria. However, it will also detect aerobic bacteria. Soybean–Casein Digest Medium is suitable for the culture of both fungi and aerobic bacteria.

下面的培养基已经被证实适合进行无菌检查。巯基醋酸盐液体培养基主要用于厌氧菌的培养。但其也用于检测好氧菌。大豆酪蛋白消化物培养基适合于培养霉菌和好氧菌。

Fluid Thioglycollate Medium 巯基醋酸盐液体培养基

L-Cystine L-胱氨酸 Sodium Chloride氯化钠 Dextrose (C6H12O6·H2O) 葡萄糖 Agar, granulated (moisture content not exceeding 15%) 0.5 g 2.5 g 5.5/5.0 g 琼脂，呈颗粒状(水分含量不超过15%) 0.75 g Yeast Extract (water-soluble) 酵母提取物(水溶性) Pancreatic Digest of Casein 酪蛋白胰酶消化物 Sodium Thioglycollate巯基乙酸钠 or Thioglycolic Acid或者巯基乙酸 Resazurin Sodium Solution (1 in 1000), freshly prepared 刃天青钠溶液(1比1000)，新配制 Purified Water 纯净水 1.0 mL 1000 mL 15.0 g 0.5 g 0.3 mL 5.0 g Mix the L-cystine, sodium chloride, dextrose, yeast extract, and pancreatic digest of casein with the purified water, and heat until solution is effected. Dissolve the sodium thioglycollate or thioglycolic acid in the solution and, if necessary, add 1 N sodium hydroxide so that, after sterilization, the solution will have a pH of 7.1 ± 0.2. If filtration is necessary, heat the solution again without boiling, and filter while hot through moistened filter paper. Add the resazurin sodium solution, mix, and place the medium in suitable vessels that provide a ratio of surface to depth of medium such that not more than the upper half of the medium has undergone a color change indicative of oxygen uptake at the end of the incubation period. Sterilize using a validated process. If the medium is stored, store at a temperature between 2 and 25 in a sterile, airtight container. If more than the upper one-third of the medium has acquired a pink color, the medium may be restored once by heating the containers in a water-bath or in free-flowing steam until the pink color disappears and by cooling quickly, taking care to prevent the introduction of nonsterile air into the container.

将L-胱氨酸、氯化钠、葡萄糖、酵母提取物、酪蛋白胰酶消化物与纯净水混合，并加热至实现溶解。将巯基乙酸钠或者巯基乙酸溶解于该溶液，如果需要可再加入1N氢氧化钠，以便在灭菌后该溶液呈pH值7.1 ± 0.2。如需要则过滤，再次加热该溶液但不得煮沸，并趁热以湿润滤纸将该溶液过滤。加入刃天青钠溶液，混匀，并将该培养基置于适当容器中，该容器应为培养基提供特定的面积－深度比，以使在培养期末表明氧气摄入的变色部分不超过培养基的上半部分。使用经过验证的工艺进行灭菌。如果需要储存该培养基，将其置于无菌、气密容器中，在2 至25 之间储藏。如果超过上部三分之一的培养基已经呈粉色，可以用以下方法恢复该培养基一次：在水浴锅中或者自由流动蒸气中加热该容器，直至粉色消失，并迅速放凉，须小心防止非无菌空气进入到容器中。

Fluid Thioglycollate Medium is to be incubated at 32.5 ± 2.5 .

巯基醋酸盐液体培养基将在32.5 ± 2.5 条件下进行培养。

Alternative Thioglycollate Medium 替代巯基醋酸盐培养基

Prepare a mixture having the same composition as that of the Fluid Thioglycollate Medium, but omitting the agar and the resazurin sodium solution, sterilize as directed above, and allow to cool prior to use. The pH after sterilization is 7.1 ± 0.2. Incubate under anaerobic conditions for the duration of the incubation period.

配制与巯基醋酸盐液体培养基成分相同，但省略了琼脂和刃天青钠溶液的混合物，按上述方法灭菌，并在使用前静置至凉。灭菌后pH值为7.1 ± 0.2。在厌氧条件下培养，培养时间同培养期。

Alternative Fluid Thioglycollate Medium is to be incubated at 32.5 ± 2.5 . 替代性巯基醋酸盐培养基将在32.5 ± 2.5 条件下进行培养。

Soybean–Casein Digest Medium 大豆-酪蛋白消化物培养基 Pancreatic Digest of Casein酪蛋白胰酶消化物 Sodium Chloride氯化钠 Dibasic Potassium Phosphate磷酸氢二钾 Dextrose (C6H12O6·H2O)葡萄糖 Purified Water纯净水 17.0 g 5.0 g 2.5 g 2.5/2.3 g 1000 mL Papaic Digest of Soybean Meal大豆粉木瓜蛋白酶消化物 3.0 g Dissolve the solids in the Purified Water, heating slightly to effect a solution. Cool the solution to room temperature, and adjust the pH with 1 N sodium hydroxide so that, after sterilization, it will have a pH of 7.3 ± 0.2. Filter, if necessary to clarify, dispense into suitable containers, and sterilize using a validated procedure. Store at a temperature between 2 and 25 in a sterile well-closed container, unless it is intended for immediate use.

将固体物质溶解于纯净水，轻微加热以实现溶解。放凉溶液至室温，并用1N氢氧化钠调整pH值，以便在灭菌后其pH值呈 7.3 ± 0.2。过滤，如需要则使之澄清，分装入适合的容器，